Human T-cell leukemia virus type 1 (HTLV-1) but not HTLV-2 is associated with adult T-cell leukemia, and the distinct pathogenicity of these two closely related viruses is thought to stem from the distinct biological functions of the respective transforming proteins, HTLV-1 Tax1 and HTLV-2 Tax2. In this study, we demonstrate that Tax1 but not Tax2 interacts with NF-kappaB2/p100 and activates it by inducing the cleavage of p100 into the active transcription factor p52. Using RNA interference methods, we further show that NF-kappaB2/p100 is required for the transformation induced by Tax1, as determined by the ability to convert a T-cell line (CTLL-2) from interleukin-2 (IL-2)-dependent to -independent growth. While Tax2 shows a reduced transforming activity relative to Tax1, Tax2 fused with a PDZ domain binding motif (PBM) present only in Tax1 shows transforming activity equivalent to that of Tax1 in CTLL-2 cells expressing an inducer of p52 processing. These results reveal that the activation of NF-kappaB2/p100 plays a crucial role in the Tax1-mediated transformation of T cells and that NF-kappaB2/p100 activation and PBM function are both responsible for the augmented transforming activity of Tax1 relative to Tax2, thus suggesting that these Tax1-specific functions play crucial roles in HTLV-1 leukemogenesis.